PROJECT SUMMARY Human astroviruses (HAstV) are a leading cause of viral diarrhea in young children. Currently, no vaccines or antiviral therapies exist for HAstV infections. Our goal is to identify sites of vulnerability (Achilles' heels) on the HAstV virion surface and exploit this information for the development of new and innovative strategies against HAstV. The HAstV virion is a small, non-enveloped icosahedral virus composed of an RNA genome surrounded by a continuous capsid protein shell studded with spikes. Despite the important role of the HAstV capsid protein in virus entry into host cells, the location of the receptor-binding site, the location of antibody epitopes, and the mechanisms of virus-neutralizing antibodies are unknown. Our central hypothesis is that the HAstV receptor-binding site is a target of HAstV-neutralizing antibodies. Using a diverse toolbox of structural, biochemical, immunological, and virological studies, we will pursue three specific aims to (1) Determine neutralizing mechanisms and serotype-reactivity of a panel of monoclonal antibodies, (2) Structurally and biophysically characterize interactions between the HAstV capsid and neutralizing monoclonal antibodies, and (3) Identify one or more HAstV capsid receptor/co-receptor binding site(s). This work represents the first molecular study of astrovirus epitopes. Results obtained by this work will elucidate mechanisms of astrovirus entry and neutralization and provide molecular blueprints for the design of vaccines and therapeutics to prevent and treat HAstV infections.